Evaluation Platform Research Articles

Multifunctional nanorobot system in precise evaluation and manipulation of virus capsid

It is imperative to have high adaptive techniques for sensing and manipulating biological targets at the nanoscale. This necessity becomes particularly crucial when dealing with fragile living bio-organisms like viruses, where the expression of capsids is closely linked to viral functions and genome constitution. Therefore, the development of a comprehensive system for dissecting and measuring viruses holds significant implications for the pharmaceutical industry and drug manufacturing. Leveraging the sub-nanometer spatial resolution and controllable tip-cantilever architecture of atomic force microscopy (AFM), a probe-laser system has been integrated as a self-sensing robotic end effector. To address intrinsic challenges in AFM-based robotic systems such as the lack of real-time monitoring, low scanning rates, and nonlinear motion caused by piezoelectric actuators, an augmented reality robotic system has been implemented. This system incorporates stereoscopic vision, a haptic feedback controller, a position recovery scheme, and a real-time force control algorithm. The integration of these components enhances the system’s capability to accurately dissect virus capsids. Operators can now perform highly efficient nanoscale tasks with multidimensional perception, utilizing the combination of stereoscopic vision and haptic force control. The position correction during manipulation can achieve a frame rate of over 30 frames per second, imperceptible to the operator, enabling closed-loop operation control. By adopting the proposed nanorobotic system in virology studies, it becomes possible to achieve accurate manipulation and dissection of SARS-CoV-2 pseudovirus capsids, and derive multi-parametric properties such as structural integrity, protein fragment thickness, and adhesive forces. The established nanobot system and experimental results serve as a guiding platform for high-accuracy evaluation in drug manufacturing development.

Towards a Remote Patient Monitoring Platform for Comprehensive Risk Evaluations for People with Diabetic Foot Ulcers

Diabetic foot ulcers (DFUs) significantly affect the lives of patients and increase the risk of hospital stays and amputation. We suggest a remote monitoring platform for better DFU care. This system uses digital health metrics (scaled from 0 to 10, where higher scores indicate a greater risk of slow healing) to provide a comprehensive overview through a visual interface. The platform features smart offloading devices that capture behavioral metrics such as offloading adherence, daily steps, and cadence. Coupled with remotely measurable frailty and phenotypic metrics, it offers an in-depth patient profile. Additional demographic data, characteristics of the wound, and clinical parameters, such as cognitive function, were integrated, contributing to a comprehensive risk factor profile. We evaluated the feasibility of this platform with 124 DFU patients over 12 weeks; 39% experienced unfavorable outcomes such as dropout, adverse events, or non-healing. Digital biomarkers were benchmarked (0–10); categorized as low, medium, and high risk for unfavorable outcomes; and visually represented using color-coded radar plots. The initial results of the case reports illustrate the value of this holistic visualization to pinpoint the underlying risk factors for unfavorable outcomes, including a high number of steps, poor adherence, and cognitive impairment. Although future studies are needed to validate the effectiveness of this visualization in personalizing care and improving wound outcomes, early results in identifying risk factors for unfavorable outcomes are promising.

Abstract PO1-23-04: The IBRAnet localisation study; a UK National multi-centre cohort study comparing the safety and effectiveness of guidewire, RFID, and magnetic seed- guided localisation for impalpable breast lesions

Abstract Background Wire-guided localisations (WGL) of breast cancers have been the gold-standard for pre-operative localisation of non-palpable breast cancers for many years but have significant limitations. Newer localisation techniques have been introduced including Radio Frequency ID tags (RFID) and magnetic seeds which aim to overcome these limitations. The UK iBRA-net group have designed a multi-centre platform study aiming to compare the safety and effectiveness of wire, RFID, magnetic seeds, and radar-based localisation. A second aim was the qualitative assessment of shared learning as users gained experience. Methods A UK national multi-centre prospective observational platform study was designed to compare a control arm of WGL for women undergoing breast conserving surgery with two interventional arms of RFID tags and magnetic seeds. Patient data were collected between August 2018 and July 2022 in UK centres. The primary outcome was accurate excision of the index lesion at the time of surgery. Secondary outcomes were defined as; resection margins, breast reoperation rate (planned and unplanned), complications, cancellation rate on day of surgery, and duration of the surgery. To account for lesion size in the assessment of excision weight, we used size as a denominator over two dimensions, reporting this as weight/size2, in g/mm2. Data was collected prospectively on a secure REDcap database following approval from local audit departments. To calculate the sample size a power calculation was employed, with n=950 patients per group the upper limit of the observed one-sided 95% confidence interval for the difference between identification rates (intervention vs WGL) is expected to be less than 0.9% with 80% power, assuming the two methods both have an expected identification rate of 99.4%. Simple summary statistics were calculated for each outcome and data were tested for distribution and differences between groups using unpaired t-tests, Mann-Whitney U tests, and Chi squared tests as appropriate. Analyses were conducted using Stata® IC version 14 (StataCorp, College Station, Texas, USA). Results Data were accrued from 3484 patients in 55 units. This included 1293 patients having WGL, 1188 RFID tags, and 1003 magnetic seeds. RFID had a lower rate of identification of the index lesion compared to magnetic seed and wire (97.9% vs. 99.8% vs. 99.1% respectively). When comparing the reasons behind failure of the primary outcome, the majority were not thought to be related to the modality of localisation. Data from 1560 patients with unifocal, unilateral breast lesions were included in subgroup analysis, excluding those having neo-adjuvant chemotherapy and those having therapeutic mammoplasty. Positive margins were lower in RFID excisions compared to wire (11.2% vs. 15% respectively, p< 0.05), whereas magnetic seed were equivalent (11.2 % vs 13.3%, P=0.22). Routine cavity shaves were performed in 64.5% of WGL vs 48.3% of RFID vs 64.3% of magnetic guided excisions. There was no difference in specimen weight/size2 (0.138 WGL vs. 0.131 RFID vs. 0.15 magnetic seed). Re-excision (for positive or close margins) was equivalent across the three groups (13.2% of wire vs 15% of RFID vs 12.3% of magnetic seed guided excisions, p=0.3). There have been four shared learning educational events focussing on three key themes relating to preoperative, intraoperative, and postoperative learning outcomes in addition to 130 database shared learning entries. This may aid other surgeons as they adopt these techniques. Conclusions All three localisation devices demonstrate high levels of localisation accuracy. There was a higher rate of device dislodgement in the RFID group, which was also identified in the shared learning events. However, re-excision rates were equivalent across the three groups. This study has demonstrated a robust platform for the comparative evaluation of new localisation technologies and of sharing learning and experience. Citation Format: Rachel Foster, James Harvey, Rajiv Dave, Anthony Maxwell, Senthurun Mylvaganam, Matthew Gardiner, Sue Down, Nicola Barnes, Mihir Chandarana, Shelley Potter, Yazan Masannat, Chris Holcombe, Tahir Masudi, Amtul Carmichael, Robert Milligan, Jenna Morgan. The IBRAnet localisation study; a UK National multi-centre cohort study comparing the safety and effectiveness of guidewire, RFID, and magnetic seed- guided localisation for impalpable breast lesions [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO1-23-04.

Abstract PO2-13-06: Analysis of Antibody Response to SV-BR-1-GM Therapeutic Vaccine in Breast Cancer Patients Using Human Protein Microarrays: Potential Correlations with Therapy Response

Abstract Background: Therapeutic cancer vaccines aim to stimulate the immune system by utilizing tumor antigens to trigger an antitumor response. In our clinical trials, we have focused on evaluating the efficacy of the breast cancer cell line secreting GM-CSF, SV-BR-1-GM as a therapeutic vaccine, and we have observed encouraging clinical outcomes. The SV-BR-1-GM regimen has been used alone (“monotherapy”, ClinicalTrials.gov NCT03066947 – study completed) and in combination with checkpoint inhibitors (“combination”, ClinicalTrials.gov NCT03328026 – study ongoing). To further improve the therapeutic efficacy of this vaccine we have initiated a study to characterize patient’s immune response to this treatment. It has been demonstrated that cancer vaccines can generate both humoral and cellular immune responses. However, predicting immune responses to cancer vaccines, especially when whole cells are employed as immunogens, presents significant challenges. We present here a preliminary analysis of the antibody response to SV-BR-1-GM in breast cancer patients using antigen arrays. Methods: Large-scale protein arrays are versatile and sensitive platforms for antibody specificity evaluation. The HuProt™ Human Proteome Microarray (CDI Laboratories, Inc., Baltimore, Maryland, United States) provides the largest number of unique, full-length, individually purified human proteins on a single microscope slide. This allows thousands of interactions to be profiled in a high-throughput manner. We utilized here an unbiased human protein microarray platform encompassing >21,000 proteins and isoforms from ~19,000 unique genes to identify IgG and IgM responses against self-antigens elicited by treatment. Serum samples from SV-BR-GM-treated patients were analyzed, comparing pre- and post-treatment. The human serum samples were probed at 1:1000 dilution. After sample processing and data collection the raw signal intensities on all arrays were quantile normalized using CDI software. Heatmaps were generated using MetaboAnalyst 5.0, which utilizes the R pheatmap package (version 0.7.7). For paired analysis, fold changes (FCs) were calculated by determining the ratio between paired pre- and post-treatment samples, resulting in one FC per pair. The means of these FCs (pair means) were then computed. ANOVA tests were performed using the RStatix R package, which automatically determines the appropriate Type I, II, or III errors for the analysis. Results: By using this approach, we were able to evaluate a broader range of antigens compared to previous investigations. Using a variety of statistical approaches, potential correlations with patient survival on SV-BR-1-GM were evaluated. Antibody responses to galectin antigens demonstrated the most consistent relationships with survival. Further confirmation with additional patients and prospective analysis are needed to fully understand relationships with clinical benefit and survival. Citation Format: Miguel Lopez-Lago, Mingjin Chang, Giuseppe Del Priore, Vikas Bhardwaj, Patience Cournoo, Pedro Ramos, Tyler Hulett, Charles Wiseman, William Williams. Analysis of Antibody Response to SV-BR-1-GM Therapeutic Vaccine in Breast Cancer Patients Using Human Protein Microarrays: Potential Correlations with Therapy Response [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO2-13-06.

Establishment of health food safety evaluation method-In vivo genotoxicity evaluation by micronucleus assay

In recent years, various factors such as shifts in health consciousness, changes in lifestyle habits, increased focus on dietary intake, the promotion of alternative medical concepts, and advancements in life sciences and technology have propelled the robust growth of the health food industry. An increasing number of reports suggest that "functional foods," containing beneficial functional components, may enhance short-term well-being and are increasingly regarded as healthful. Building upon these findings, this study aims to enhance the production capacity of high-value agricultural products. However, for quality assurance, toxicity assessment of functional foods is imperative. There is a growing necessity to employ in vivo genotoxicity assays to evaluate the carcinogenic potential of these products. In this study, we tried to establish another in vivo genotoxicity evaluation platform via acridine orange induction. According to all results in this study, during the experiment, there were no statistically significant differences in BW between the negative control group and the normal control group (p > 0.05). Moreover, the clinical observations of the experimental mice showed that all mice survived until the end of the experiment, and no abnormal clinical symptoms were observed. The mouse’s food consumption in each group were monitored during the experiments. The food consumption of mice was recorded daily after the experiment. During the experiments, there were no significant difference between two groups on the food consumption. The percentage of RETs/1,000 RBCs in the negative control group was significantly decrease than the normal control group (p < 0.001). RETs/1,000 RBCs (‰) in ICR mice (n = 5/group) in the negative control group and the normal control group at 48th and 72th hours-experiment were respectively 18 ± 4.6 / 45.6 ± 4.0 (48th hours-experiment) and 19 ± 4.8 / 47.6 ± 4.8 (72th hours-experiment). The percentage of Mn-RETs/1,000 RETs in the negative control group was significantly increase than the normal control group (p < 0.001). Mn-RETs/1,000 RETs (‰) in ICR mice (n = 5/group) in the negative control group and the normal control group were 10.6 ± 2.1 / 0.4 ± 0.5 and 8.2 ± 1.8 / 0.4 ± 0.5, respectively. Taken all results of RET/RBCs (‰) and MN-RET/RETs (‰) together, the genotoxicity in the negative control group has been successfully induced. In light of these requirements, we propose utilizing the mammalian erythrocyte micronucleus assay to detect damage induced by test substances to erythroblast chromosomes. This study endeavors to establish a sensitive and stable method for quantifying micronuclei formation in erythrocytes in vivo. The resulting data will facilitate the evaluation of potential toxicity associated with agricultural functional products in the future.

A vascularized breast cancer spheroid platform for the ranked evaluation of tumor microenvironment-targeted drugs by light sheet fluorescence microscopy

Targeting the supportive tumor microenvironment (TME) is an approach of high interest in cancer drug development. However, assessing TME-targeted drug candidates presents a unique set of challenges. We develop a comprehensive screening platform that allows monitoring, quantifying, and ranking drug-induced effects in self-organizing, vascularized tumor spheroids (VTSs). The confrontation of four human-derived cell populations makes it possible to recreate and study complex changes in TME composition and cell-cell interaction. The platform is modular and adaptable for tumor entity or genetic manipulation. Treatment effects are recorded by light sheet fluorescence microscopy and translated by an advanced image analysis routine in processable multi-parametric datasets. The system proved to be robust, with strong interassay reliability. We demonstrate the platform’s utility for evaluating TME-targeted antifibrotic and antiangiogenic drugs side-by-side. The platform’s output enabled the differential evaluation of even closely related drug candidates according to projected therapeutic needs.

Multiplex analysis platform for evaluation of endocrine disruption of emerging contaminants against human steroid hormone receptors using autobioluminescent yeast bioassay: Application to bisphenols

Emerging synthetic or derivative chemicals have drawn global concern due to their potential endocrine-disrupting effects, either as agonists or antagonists or through indirect effects on enzymes of signal transduction pathways. Currently, in vitro screening models are limited to poor tolerance, robustness, high cost and need for sphisticated equipment or complex procedures that require high levels of expertise. Herein, a multiplex analysis platform based on autobioluminescent yeast strains, including BLYhERαS, BLYhERβS, BLYhPRS, BLYhARS, BLYhMRS, and BLYhGRS, was developed for use in easy, rapid, robust, and sensitive screening for potential modulation of endocrine systems. Methods were applied to assess endocrine-disrupting effects of Bisphenol A (BPA) and its alternatives. Disrupting activities of bisphenols (BPs) varied greatly, and some BPA substitutes exerted more potent activities than those of BPA. Most BPs were primarily agonists of ERα and ERβ, while others were antagonists to AR, PR, GR, and MR. BPP and BPM exhibited non-monotonic dose-response relationships toward ERα and ERβ, and BHPF and BPG displayed converse disrupting activities to ERα and ERβ. Overall, the bioassay system provides an efficient tool for comprehensive evaluation of endocrine-disrupting activities of emerging contaminants.

BuildingTwin - Open Platform for Monitoring, Evaluation and Optimization of Building Operation

BuildingTwin - Open Platform for Monitoring, Evaluation and Optimization of Building Operation

Ultrasensitive single-step CRISPR detection of monkeypox virus in minutes with a vest-pocket diagnostic device

The emerging monkeypox virus (MPXV) has raised global health concern, thereby highlighting the need for rapid, sensitive, and easy-to-use diagnostics. Here, we develop a single-step CRISPR-based diagnostic platform, termed SCOPE (Streamlined CRISPR On Pod Evaluation platform), for field-deployable ultrasensitive detection of MPXV in resource-limited settings. The viral nucleic acids are rapidly released from the rash fluid swab, oral swab, saliva, and urine samples in 2 min via a streamlined viral lysis protocol, followed by a 10-min single-step recombinase polymerase amplification (RPA)-CRISPR/Cas13a reaction. A pod-shaped vest-pocket analysis device achieves the whole process for reaction execution, signal acquisition, and result interpretation. SCOPE can detect as low as 0.5 copies/µL (2.5 copies/reaction) of MPXV within 15 min from the sample input to the answer. We validate the developed assay on 102 clinical samples from male patients / volunteers, and the testing results are 100% concordant with the real-time PCR. SCOPE achieves a single-molecular level sensitivity in minutes with a simplified procedure performed on a miniaturized wireless device, which is expected to spur substantial progress to enable the practice application of CRISPR-based diagnostics techniques in a point-of-care setting.

Performance Evaluation of an Enhanced Solar Still Using Simulink

This study focuses on enhancing the efficiency of solar desalination, particularly for small communities facing water scarcity. Despite the economic viability of solar desalination, the method encounters a technical challenge related to low solar yield. The research proposes innovative solutions, notably the integration of a Compound Parabolic Collector (CPC) and Thermal Energy Storage (TES). The CPC system addresses low solar yield by minimizing losses through reflection, significantly improving the capture efficiency of solar radiation. Additionally, TES serves as a strategic storage component, storing solar energy and providing high energy when needed, thereby enhancing water evaporation rates. Using the SIMULINK platform for evaluation, the study demonstrates that the solar still, enhanced with TES and CPC, consistently produces an average of 15 liters of clean water. This positive outcome underscores the effectiveness of the proposed enhancements and suggests avenues for further refinement. The research not only addresses technical challenges but also establishes a foundation for ongoing improvements in solar desalination technology, offering potential solutions for water-scarce communities.

Total Synthesis of Core 3 & Core 4‐Type Mucin Glycan Derivatives

Recent studies have demonstrated the ability of mucin O‐glycans to attenuate virulence in diverse, cross‐kingdom pathogens, sparking interest in their development as a novel therapeutic approach against infection. Although their virulence attenuating activity is evident, mucin glycans obtained from native sources comprise mixtures of several hundred distinct structures, and therefore the specific active glycan epitopes and molecular mechanisms of virulence attenuation remain unclear. Individual mucin glycan structures cannot be purified from native sources and are not amenable to automated synthesis; therefore, to further investigate the phenomena of virulence attenuation we have been developing convergent and scalable methods (>30 mg per target glycan) to assemble a mucin O‐glycan library in sufficient scale and purity to facilitate biological studies. Previously we described a method to obtain core 1 & core 2‐type mucin glycan derivatives that have since been used to identify active epitopes in Candida albicans and Vibrio cholerae. In the current study, we describe the expansion of our glycan library to include core 3 & core 4‐type derivatives, increasing the structural diversity of our platform for biological evaluation.

180 Building an evaluation platform to capture the impact of Frontiers CTSI activities

OBJECTIVES/GOALS: In 2021, Frontiers CTSI revamped its evaluation infrastructure to be comprehensive, efficient, and transparent in demonstrating outputs and outcomes. We sought to build a platform to standardize measures across program areas, integrate continuous improvement processes into operations, and reduce the data entry burden for investigators. METHODS/STUDY POPULATION: To identify useful metrics, we facilitated each Core’s creation of a logic model, in which they identified all planned activities, expected outputs, and anticipated outcomes for the 5-year cycle and beyond. We identified appropriate metrics based on the logic models and aligned metrics across programs against extant administrative data. We then built a data collection and evaluation platform within REDCap to capture user requests, staff completion of requests, and, ultimately, request outcomes. We built a similar system to track events, attendance, and outcomes. Aligning with other hubs, we also transitioned to a membership model. Membership serves as the backbone of the evaluation platform and allows us to tailor communication, capture demographic information, and reduce the data entry burden for members. RESULTS/ANTICIPATED RESULTS: The Frontiers Evaluation Platform consists of 9 redcap projects with distinct functions and uses throughout the Institute. Point-of-service collection forms include the Consultation Request Event Tracking. Annual Forms include a Study Outcome, Impact, and Member Assessment Survey. Set timepoint collections include K & T application, Mock Study Section, and Pilot grant application submission, review, and outcomes. Flight Tracker is used to collect scientific outcomes and integrated with the platform. Using SQL, the membership module has been integrated into all forms to check and collect membership before service access and provide relevant member data to navigators. All relevant data is then synched into a dashboard for program leadership and management to track outputs and outcomes in real-time. DISCUSSION/SIGNIFICANCE: Since the launch of the evaluation platform in Fall 2022, Frontiers has increased its workflow efficiency and streamlined continuous improvement communication. The platform can serve as a template for other hubs to build efficient processes to create comprehensive and transparent evaluation plans.

152 Building the Future of Dissemination and Implementation (D&I) Science at Frontiers CTSI: Capacity Building, Infrastructure, and Emerging Research Areas

OBJECTIVES/GOALS: A diverse workforce is an aspiration of CTSIs and embedded in goals to build D&I workforce capacity. However, little research describes the diversity of the current workforce. We assessed current assets, opportunities, and diversity of D&I efforts at Frontiers and characterized investigators and those supported by services offered. METHODS/STUDY POPULATION: In January 2021, Frontiers convened a working group to identify existing D&I assets and needs in the CTSI multi-state catchment area. The committee catalogued existing training and consultation resources and services, which the CTSI supported with infrastructure to support, track, and evaluate ongoing efforts. We obtained data from the evaluation platform and conducted descriptive analyses of the investigators and service uptake among two programs offered, contrasting the workforce with national data obtained from the American Academy of Medical Colleges (2022) and the National Institutes of Health (2018). RESULTS/ANTICIPATED RESULTS: Ninety individuals at 9 institutions across Kansas and Missouri identified as implementation researchers. Since 2022, 28 D&I consultations were provided, 92% for grant applications. Five early-stage investigators were identified for career development in an NIH-supported Health Equity and Implementation Center (ESI). The network mirrors the larger workforce regarding underrepresented racial/ethnic minorities (18%) and new investigators (60%). More women (76%) are represented in the D&I network and among ESIs (80%) than the academic workforce (44%), but significantly fewer women used consultation services (p<0.001). Lower proportions of underrepresented minority investigators (p<0.001) and investigators from disadvantaged statuses (p=0.027) accessed consultations services. DISCUSSION/SIGNIFICANCE: Investigators underrepresented in science on multiple dimensions were less likely to use consulting services, Outreach for consultation services may be necessary, if needs are not being met in other programs. Further exploration of overall D&I workforce trends is needed to ensure goals for the field and the CTSA network are achieved.

Visual screening of CRISPR/Cas9 editing efficiency based on micropattern arrays for editing porcine cells.

CRISPR/Cas9 technology, combined with somatic cell nuclear transplantation (SCNT), represents the primary approach to generating gene-edited pigs. The inefficiency in acquiring gene-edited nuclear donors is attributed to low editing and delivery efficiency, both closely linked to the selection of CRISPR/Cas9 forms. However, there is currently no direct method to evaluate the efficiency of CRISPR/Cas9 editing in porcine genomes. A platform based on fluorescence reporting signals and micropattern arrays was developed in this study, to visually assess the efficiency of gene editing. The optimal specifications for culturing porcine cells, determined by the quantity and state of cells grown on micropattern arrays, were a diameter of 200µm and a spacing of 150µm. By visualizing the area of fluorescence loss and measuring the gray value of the micropattern arrays, it was quickly determined that the mRNA form targeting porcine cells exhibited the highest editing efficiency compared to DNA and Ribonucleoprotein (RNP) forms of CRISPR/Cas9. Subsequently, four homozygotes of the β4GalNT2 gene knockout were successfully obtained through the mRNA form, laying the groundwork for the subsequent generation of gene-edited pigs. This platform facilitates a quick, simple, and effective evaluation of gene knockout efficiency. Additionally, it holds significant potential for swiftly testing novel gene editing tools, assessing delivery methods, and tailoring evaluation platforms for various cell types.

Modeling and Subjective Evaluation Method of Driveability for Fuel Cell Vehicles

Aiming at the demand for subjective evaluation of driveability for fuel cell vehicles, the modeling and evaluation method of driveability for fuel cell vehicles were studied in this paper. Firstly, a real-time model of the fuel cell vehicle powertrain system was established, which included the fuel cell model, power battery model, DC/DC converter model and drive motor model. Secondly, it was integrated with the vehicle dynamics model to form a virtual prototype of a fuel cell vehicle. And a virtual subjective evaluation platform for fuel cell vehicles was built by combining the virtual prototype and high-fidelity driving simulator. Thirdly, a subjective evaluation method of driveability for fuel cell vehicles was proposed, which included the starting performance, acceleration performance, uniform speed performance and tip-in/tip-out performance. Finally, the paper used the platform and method mentioned above to conduct subjective evaluations of the fuel cell vehicles.

Abstract 5345: Development and validation of humanized HLA mouse model platforms for preclinical evaluation of novel peptide vaccines

Abstract The human leukocyte antigen (HLA) class I major histocompatibility complex (MHC) plays an integral role in immune surveillance by presenting intracellular antigen peptides on the cell surface, enabling recognition by cytotoxic T cells and subsequent elimination. Harnessing this intrinsic process for cancer immunotherapy is a promising therapeutic strategy; thus, developing preclinical models to evaluate the efficacy of novel peptide vaccines in vivo and ex vivo is critical. Previously, we developed a humanized MHC I mouse model (B-HLA-A2.1 mice), in which we successfully demonstrated efficacy of peptide vaccines as assessed by splenocyte IFN-γ production measured via ELISpot. Here, we provide evidence that B-HLA-A2.1 mice can also mount immune responses to NY-ESO-1 and WT1-Db126 peptides using the same assay. Additionally, immunization with WT1-Db126 at 7, 14, and 21 days prior to tumor inoculation with WT1-expressing MC38 colon cancer cells significantly reduced tumor volume. Additionally, we have generated two new humanized HLA models: B-HLA-A11.1 mice and B-HLA-A24.2 mice. Flow cytometric data demonstrate successful expression of the human HLAs on the cell surface of splenocytes isolated from both strains. Compared to wild-type C57BL/6 mice, analysis of leukocyte and lymphocyte subpopulations in the humanized mice indicates that the CD4/CD8 ratio is altered in favor of CD4+ T cells in both strains, similar to what was observed in B-HLA-A2.1 humanized mice. Despite these observed alterations, B-HLA-A24.2 mice were able to mount an immune response to peptide SART2 93-101, which was measured by splenocyte IFN-γ production. In summary, our data indicate that humanized HLA-A mice provide a powerful preclinical model for in vivo/ex vivo evaluation of novel peptide vaccines. Citation Format: Zhenlan Niu, Xuan Yu, Zhiyuan Shen, Jing Guo, Qingcong Lin, Hui Xu. Development and validation of humanized HLA mouse model platforms for preclinical evaluation of novel peptide vaccines [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5345.

Abstract 4190: Patient-derived xenograft (PDX) models with differential HER2 expression for preclinical evaluation of HER2-targeted therapies

Abstract Human epidermal growth factor receptor 2 (HER2) is a critical therapeutic target in gastric cancer. However, the varying expression levels of HER2 in patients present a significant challenge in selecting the appropriate HER2-targeted therapies. Patient-derived xenograft (PDX) models offer a valuable tool to address this clinical concern. We established a panel of PDX models derived from gastric cancer patients, each representing a distinct range of HER2 expression levels. These models were subsequently treated with HER2-targeted drugs, including Trastuzumab (Herceptin), Margetuximab, and Enhertu. Our results demonstrated differential drug responses corresponding to the varying HER2 expression levels in the PDX samples. Trastuzumab, Margetuximab, and Enhertu exhibited distinct efficacies across the HER2 expression spectrum. This underscores the significance of selecting the right therapeutic approach based on the HER2 status of individual patient. Our collection of PDX models with differential HER2 expression levels provides a robust platform for preclinical evaluation of HER2-targeted therapies. This approach not only addresses the clinical challenge of HER2 heterogeneity but also facilitates a more personalized and effective treatment strategy for gastric cancer patients. Citation Format: Xiaoliu Yang, Shiying Guo, Huiyi Wang, Xu Su, Jing Zhao, Xiang Gao, Cunxiang Ju. Patient-derived xenograft (PDX) models with differential HER2 expression for preclinical evaluation of HER2-targeted therapies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4190.

Abstract 6897: Defining critical features for successful preclinical use of glioblastoma patient-derived xenografts (PDX) models for therapeutic assessment and clinical trial development

Abstract Mayo Clinic has developed 111 GBM PDXs from newly diagnosed (72 PDX) and recurrent (39 PDX) patients. This large collection of PDXs provides a broad representation of genetic heterogeneity seen in patients. GBM PDXs that perform well in both in vitro and in vivo settings can be extensively utilized in the preclinical setting for downstream analysis. Based on 20 years of experience, we have defined the attributes that best enable preclinical use of our GBM PDX models. Among them are short-term in vitro culturing performance, large scale genomic evaluation data, benchmarked responses to standards of care, and median time to moribund for untreated orthotopic tumors. PDXs with prolonged survival timelines delay study readouts and, in some situations, confound efficacy results due to age-related animal decline. Setting median survival thresholds as part of the upfront experimental design allows for more timely study readouts that are unaffected by these confounding variables. Our preferred cutoff for the Mayo GBM PDX collection is 70 days median survival (n=64). Using this 64 PDX line panel, regimens expected to double survival would produce results within 4-5 months. This timeframe is well within the range of robust health for athymic nude mice. This 64-line collection features both newly diagnosed (nGBM; n=42) and recurrent (rGBM; n=22) tumors. MGMT promoter methylation status was determined by qMS-PCR (25 methylated, 37 unmethylated, 2 indeterminate). Available WES (n=62) and RNAseq (n=42) data, along with our tissue microarray (n=42), are critical resources for biomarker-based evaluations aimed at defining patient populations that are most likely to benefit from a given therapeutic. From these data, the molecular subtypes available include 26 classical, 14 proneural, and 3 mesenchymal. Twenty-seven of these 64 PDX models have also been successfully cultured from fresh as well as cryopreserved cell stocks to further expand their experimental value. Correlative clinical data including diagnosis and treatments are also available (n=61). Current standards of care for GBM include radiation, temozolomide, and bevacizumab. Defining sensitivity to these therapies is an important first step for evaluating the integration of novel therapeutics with these standards. Since the creation of our GBM PDX collection, efficacy of these therapies has been evaluated as follows: RT 26 PDXs, TMZ 20 PDXs, and Avastin 23 PDXs. The Mayo Clinic GBM PDX collection provides an important platform for preclinical evaluation of novel therapeutics. Through careful selection of PDX lines with favorable growth characteristics and known genetic features, researchers can select the most relevant models for their study. This approach will enable more meaningful preclinical work in the hopes of providing more tangible clinical benefits for GBM patients. Citation Format: Danielle M. Burgenske, Ann C. Mladek, Katrina K. Bakken, Zeng Hu, Brett L. Carlson, Paul A. Decker, Jeanette E. Eckel-Passow, Jann N. Sarkaria. Defining critical features for successful preclinical use of glioblastoma patient-derived xenografts (PDX) models for therapeutic assessment and clinical trial development [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6897.

Abstract 2831: Development and application of the NCG-hIL2 mouse model: A humanized platform for enhanced NK cell evaluation in ADCC efficacy testing

Abstract Natural Killer (NK) cells play a pivotal role in immune surveillance and antitumor responses, making them valuable targets in cancer immunotherapy. The assessment of therapeutic antibodies' efficacy, particularly in the context of Antibody-Dependent Cell-Mediated Cytotoxicity (ADCC), necessitates models that faithfully recapitulate human NK cell responses. Here, we introduce the NCG-hIL2 mouse model, designed to address these requirements and advance our understanding of NK cell development. The NCG-hIL2 mouse model was engineered to express human Interleukin-2 (IL2) cytokine, a critical factor in NK cell development and activation. This humanized platform allows for the robust reconstitution of human NK cells through the transplantation of peripheral blood mononuclear cells (PBMCs) or hematopoietic stem cells (HSCs). In this study, we validate the NCG-hIL2 model's exceptional capability in supporting the reconstitution of human NK cells, highlighting the indispensable role of IL2 in NK cell development. Moreover, we demonstrate its utility in the evaluation of therapeutic antibodies, specifically Trastuzumab and the ADCC-enhanced Margetuximab, in the context of ADCC efficacy testing. Our findings reveal that the NCG-hIL2 model provides a unique advantage for assessing the impact of IL2 on NK cell development, thus facilitating a deeper understanding of the complex interplay between NK cells and therapeutic antibodies. The NCG-hIL2 mouse model serves as a valuable tool for studying NK cell development and evaluating the efficacy of immunotherapies, particularly in ADCC. By reconstituting human NK cells and leveraging the role of IL2, this model offers an enhanced platform for preclinical evaluation of therapeutic antibodies, ultimately advancing our ability to harness NK cell-mediated antitumor responses. It is poised to contribute significantly to the field of cancer immunotherapy and drug development. Citation Format: Yuan Fang, Hongyan Sun, Huiyi Wang, Jun Xing, Jialu Fan, Jing Zhao, Xiang Gao, Cunxiang Ju. Development and application of the NCG-hIL2 mouse model: A humanized platform for enhanced NK cell evaluation in ADCC efficacy testing [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2831.

Performance Evaluation of Cable Shaft Fireproof Sealing System in High-Rise Buildings: A Comparative Test Method

The effectiveness of fireproof sealing systems in preventing the spread of fire in high-rise building cable shafts relies on the properties of various sealing materials and the construction process. Therefore, a comprehensive evaluation is necessary. The authors of this paper propose a comparative test method based on an entity test platform for a performance evaluation of cable shaft fireproof sealing systems in high-rise buildings. The test platform measures changes in temperature, humidity, and smoke mass during fire tests to compare the performance of four sets of fireproof sealing systems in terms of thermal insulation, smoke sealing capacity, and overall integrity. In addition, a fire dynamics simulation (FDS) of fireproof sealing systems was carried out on the entity test platform, and the sealing failure process in the case of cracking in the fireproof sealing system was revealed. The simulation results for the temperature trends in the lower space align with the fire test results. Furthermore, as the gap size increases, the diffusion of smoke and flame accelerates. Consequently, the performance of cable shaft fireproof sealing systems depends not only on the sealing material but also on the construction process.